New GJA8 Variants and Phenotypes Highlight Its Critical Role in a Broad Spectrum of Eye Anomalies
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Human Genetics (2019) 138:1027–1042 https://doi.org/10.1007/s00439-018-1875-2 ORIGINAL INVESTIGATION New GJA8 variants and phenotypes highlight its critical role in a broad spectrum of eye anomalies Fabiola Ceroni1 · Domingo Aguilera‑Garcia2,3 · Nicolas Chassaing4,5 · Dorine Arjanne Bax1 · Fiona Blanco‑Kelly2,3,6,7 · Patricia Ramos2,3 · Maria Tarilonte2,3 · Cristina Villaverde2,3 · Luciana Rodrigues Jacy da Silva2,3 · Maria Juliana Ballesta‑Martínez8 · Maria Jose Sanchez‑Soler8 · Richard James Holt1 · Lisa Cooper‑Charles9 · Jonathan Bruty9 · Yvonne Wallis9 · Dominic McMullan9 · Jonathan Hofman10 · David Bunyan11 · Alison Stewart12 · Helen Stewart6 · Katherine Lachlan13,14 · DDD Study15 · Alan Fryer16 · Victoria McKay16 · Joëlle Roume17 · Pascal Dureau18 · Anand Saggar19 · Michael Grifths9 · Patrick Calvas4,5 · Carmen Ayuso2,3 · Marta Corton2,3 · Nicola K Ragge1,10 Received: 22 November 2017 / Accepted: 9 February 2018 / Published online: 20 February 2018 © Springer-Verlag GmbH Germany, part of Springer Nature 2018 Abstract GJA8 encodes connexin 50 (Cx50), a transmembrane protein involved in the formation of lens gap junctions. GJA8 muta- tions have been linked to early onset cataracts in humans and animal models. In mice, missense mutations and homozygous Gja8 deletions lead to smaller lenses and microphthalmia in addition to cataract, suggesting that Gja8 may play a role in both lens development and ocular growth. Following screening of GJA8 in a cohort of 426 individuals with severe con- genital eye anomalies, primarily anophthalmia, microphthalmia and coloboma, we identifed four known [p.(Thr39Arg), p.(Trp45Leu), p.(Asp51Asn), and p.(Gly94Arg)] and two novel [p.(Phe70Leu) and p.(Val97Gly)] likely pathogenic variants in seven families. Five of these co-segregated with cataracts and microphthalmia, whereas the variant p.(Gly94Arg) was identifed in an individual with congenital aphakia, sclerocornea, microphthalmia and coloboma. Four missense variants of unknown or unlikely clinical signifcance were also identifed. Furthermore, the screening of GJA8 structural variants in a subgroup of 188 individuals identifed heterozygous 1q21 microdeletions in fve families with coloboma and other ocular and/or extraocular fndings. However, the exact genotype–phenotype correlation of these structural variants remains to be established. Our data expand the spectrum of GJA8 variants and associated phenotypes, confrming the importance of this gene in early eye development. Introduction anomalies, such as early onset cataract and anterior segment dysgenesis (ASD). They are associated with extraocular fea- Anophthalmia (absent eye), microphthalmia (small eye) tures in just over half of cases (Shah et al. 2012) and can and coloboma (optic fssure closure defects), collectively form part of a syndrome (Slavotinek 2011). The etiology referred to as AMC, form a spectrum of developmental of AMC is characterised by marked genetic heterogeneity. eye disorders, with an overall estimated incidence of 6–13 This refects the complexity underlying eye morphogenesis, per 100,000 births (Shah et al. 2011; Skalicky et al. 2013). a conserved process that requires a series of highly coordi- AMC can occur alone or in combination with other ocular nated events, both at the molecular and the structural level, and is tightly regulated by a network of transcription factors, extracellular signaling molecules, cell-cycle regulators, and Marta Corton and Nicola Ragge: joint senior authors. adhesion proteins (Reis and Semina 2015). Electronic supplementary material The online version of this Connexins (Cxs) are a homogeneous family of transmem- article (https://doi.org/10.1007/s00439-018-1875-2) contains brane proteins with a crucial role in intercellular communi- supplementary material, which is available to authorized users. cation. They present a conserved topology, which consists of four transmembrane α-helices (TM1–TM4) joined by * Nicola K Ragge [email protected]; [email protected] two extracellular loops (ECL1 and ECL2) and one cytoplas- mic loop (ICL), fanked by a short cytoplasmic N-terminal Extended author information available on the last page of the article Vol.:(0123456789)1 3 1028 Human Genetics (2019) 138:1027–1042 domain (NT) and a long cytoplasmic and less conserved (Chung et al. 2007), indicating that any signifcant dysregu- C-terminal domain (CT). Cxs oligomerise in hexameric lation of Gja8 could be deleterious for eye development. complexes called connexons, and allow the transmembrane In humans, missense and frameshift mutations in GJA8 passage of ions and small solutes (≤ 1 kDa). Connexons can (OMIM 600897) have been associated with cataracts (Beyer function independently as hemichannels (HCs) or they can et al. 2013; Yu et al. 2016). Rarely, the phenotype also dock with their counterparts on the juxtaposed cell to form includes additional ocular abnormalities, mainly micro- a gap junction channel (GJC), enabling the direct exchange cornea and iris hypoplasia (Devi and Vijayalakshmi 2006; of small molecules. Given their role in cell–cell communica- Hansen et al. 2007; Hu et al. 2010; Sun et al. 2011; Prokudin tion and tissue homeostasis, Cxs have been implicated in a et al. 2014; Ma et al. 2016), but in a few cases also microph- variety of biological and pathological processes (Pfenniger thalmia (Ma et al. 2016) and sclerocornea (Ma et al. 2018). et al. 2011; García et al. 2016), including myelin-related Interestingly, defects in the formation of the lens have also diseases (Cx32 and Cx47), heart malformations and arrhyth- been observed (Ma et al. 2018). The cataracts described in mia (Cx40), hearing loss and skin disorders (Cx26, Cx30, these individuals vary in both their location (e.g., nuclear, Cx30.3, and Cx31), oculodentodigital dysplasia, a syndrome zonular, lamellar, or total) and appearance (e.g., total, pul- also involving microphthalmia, microcornea, cataract and/ verulent, or dense). The mutations are predominantly het- or spherophakia (Cx43), and early onset cataract (Cx46 and erozygous and only few homozygous variants have been Cx50). reported, all in consanguineous families (Ponnam et al. As with AMC, developmental or early onset cataracts are 2007; Schmidt et al. 2008 Ponnam et al. 2013; Ma et al. a clinically heterogeneous group of disorders, presenting as 2016). These pathogenic variants lead to amino acid altera- isolated anomalies or part of a syndrome. More than 110 tions distributed throughout the protein (Yu et al. 2016), genes have been implicated in congenital cataracts (Gillespie although mostly localised between the domains TM1 and et al. 2014), with mutations in Cxs accounting for around TM2. They are predicted to afect protein function through 16% of cases with a known genetic cause (Shiels and Hejt- various mechanisms, such as by inducing misfolding and/ mancik 2017). Since the lens does not have any blood sup- or mislocalisation or by altering channel properties (Beyer ply, it strongly depends on an extensive network of GJCs et al. 2013). for the intercellular communications that are critical for its Copy number variants (CNVs) in the distal region of development and the maintenance of its transparency. The chromosome 1q21 and including GJA8 are rare in the gen- most abundant Cxs in the lens are Cx46 and Cx50, which eral population, but have recurrently been identifed in indi- can also form mixed hexamers. Cx46, encoded by GJA3, viduals with a broad range of diferent clinical diagnoses is expressed only in fber cells, whereas Cx50, encoded by (Brunetti-Pierri et al. 2008; Meford et al. 2008; Stefansson GJA8, is present throughout the lens. et al. 2008; Bernier et al. 2016). These primarily include Genetic studies in mice have demonstrated that the developmental delay, microcephaly and psychiatric disor- homozygous knockout of either Gja3 or Gja8 leads to cata- ders, although the enrichment of 1q21 CNVs in individuals racts, but with important phenotypic diferences. The dele- with these disorders could partly be related to ascertain- tion of Gja3 causes severe progressive nuclear cataracts, but ment bias. However, some cases also have eye anomalies, does not alter ocular growth (Gong et al. 1997). In contrast, such as cataracts (Brunetti-Pierri et al. 2008; Meford et al. Gja8-null mice develop milder nuclear cataracts at an early 2008; Rosenfeld et al. 2012; Bernier et al. 2016; Ha et al. postnatal age and exhibit signifcantly smaller lenses and 2016), microphthalmia (Meford et al. 2008), and coloboma microphthalmia (White et al. 1998; Rong et al. 2002), indi- (Brunetti-Pierri et al. 2008). Among the genes afected by cating that the two Cxs have overlapping, but distinct func- these recurrent microdeletions/microduplications, GJA8 rep- tions. In addition, the targeted replacement of Gja8 with resents a strong candidate for the ocular anomalies described Gja3 (Cx50KI46 knockin mice) prevents the loss of crys- in some of the 1q21 CNV carriers. talline solubility, but not the postnatal growth defect result- To investigate further the importance of GJA8 in human ing from the Gja8 deletion (White 2002), confrming the eye morphogenesis and provide a better understanding of the functional diversity of the two proteins and the involvement range of developmental ocular anomalies associated with of Gja8 in the control of normal ocular growth. This is also mutations in this gene, we screened GJA8 in a cohort of 426 supported by mouse lines carrying missense mutations in unrelated patients (304 UK, 121 Spanish and